US4743549A - Hydrogen peroxide-forming sarcosine oxidase - Google Patents
Hydrogen peroxide-forming sarcosine oxidase Download PDFInfo
- Publication number
- US4743549A US4743549A US06/868,262 US86826286A US4743549A US 4743549 A US4743549 A US 4743549A US 86826286 A US86826286 A US 86826286A US 4743549 A US4743549 A US 4743549A
- Authority
- US
- United States
- Prior art keywords
- sarcosine
- sarcosine oxidase
- chainia
- enzyme
- oxidase
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Lifetime
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- 108010060059 Sarcosine Oxidase Proteins 0.000 title claims abstract description 27
- 102000008118 Sarcosine oxidase Human genes 0.000 title claims abstract description 27
- 229910052739 hydrogen Inorganic materials 0.000 title claims abstract 3
- 239000001257 hydrogen Substances 0.000 title claims abstract 3
- 125000004435 hydrogen atom Chemical class [H]* 0.000 title claims abstract 3
- LWIHDJKSTIGBAC-UHFFFAOYSA-K tripotassium phosphate Chemical compound [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 claims abstract description 14
- 230000000694 effects Effects 0.000 claims abstract description 8
- 229910000160 potassium phosphate Inorganic materials 0.000 claims abstract description 7
- 235000011009 potassium phosphates Nutrition 0.000 claims abstract description 7
- 241000204060 Streptomycetaceae Species 0.000 claims abstract description 6
- 239000013543 active substance Substances 0.000 claims abstract description 3
- 230000000977 initiatory effect Effects 0.000 claims abstract description 3
- 102000004190 Enzymes Human genes 0.000 claims description 24
- 108090000790 Enzymes Proteins 0.000 claims description 24
- 241000187747 Streptomyces Species 0.000 claims description 12
- 241000970890 Streptomyces purpurogeneiscleroticus Species 0.000 claims description 9
- 241000946902 Streptomyces flocculus Species 0.000 claims description 5
- 238000000034 method Methods 0.000 claims description 4
- 241000187095 Streptomyces purpureus Species 0.000 claims description 3
- 241000894007 species Species 0.000 claims description 3
- 238000012258 culturing Methods 0.000 claims description 2
- 239000002028 Biomass Substances 0.000 claims 1
- DDRJAANPRJIHGJ-UHFFFAOYSA-N creatinine Chemical compound CN1CC(=O)NC1=N DDRJAANPRJIHGJ-UHFFFAOYSA-N 0.000 description 30
- FSYKKLYZXJSNPZ-UHFFFAOYSA-N sarcosine Chemical compound C[NH2+]CC([O-])=O FSYKKLYZXJSNPZ-UHFFFAOYSA-N 0.000 description 30
- 229940088598 enzyme Drugs 0.000 description 23
- 239000003153 chemical reaction reagent Substances 0.000 description 21
- JOCBASBOOFNAJA-UHFFFAOYSA-N N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid Chemical compound OCC(CO)(CO)NCCS(O)(=O)=O JOCBASBOOFNAJA-UHFFFAOYSA-N 0.000 description 15
- 108010077895 Sarcosine Proteins 0.000 description 15
- 239000007994 TES buffer Substances 0.000 description 15
- 229940109239 creatinine Drugs 0.000 description 15
- 229940043230 sarcosine Drugs 0.000 description 15
- 239000000287 crude extract Substances 0.000 description 11
- 229910019142 PO4 Inorganic materials 0.000 description 10
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 10
- 239000010452 phosphate Substances 0.000 description 10
- 238000006243 chemical reaction Methods 0.000 description 9
- 239000006228 supernatant Substances 0.000 description 9
- RLFWWDJHLFCNIJ-UHFFFAOYSA-N 4-aminoantipyrine Chemical compound CN1C(C)=C(N)C(=O)N1C1=CC=CC=C1 RLFWWDJHLFCNIJ-UHFFFAOYSA-N 0.000 description 8
- CVSVTCORWBXHQV-UHFFFAOYSA-N creatine Chemical compound NC(=[NH2+])N(C)CC([O-])=O CVSVTCORWBXHQV-UHFFFAOYSA-N 0.000 description 8
- 230000002255 enzymatic effect Effects 0.000 description 8
- 239000000523 sample Substances 0.000 description 8
- 239000003599 detergent Substances 0.000 description 7
- 241000194110 Bacillus sp. (in: Bacteria) Species 0.000 description 6
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 6
- 238000004458 analytical method Methods 0.000 description 6
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 4
- 229960003624 creatine Drugs 0.000 description 4
- 239000006046 creatine Substances 0.000 description 4
- 239000008363 phosphate buffer Substances 0.000 description 4
- 102000004169 proteins and genes Human genes 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- NRHMKIHPTBHXPF-TUJRSCDTSA-M sodium cholate Chemical compound [Na+].C([C@H]1C[C@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC([O-])=O)C)[C@@]2(C)[C@@H](O)C1 NRHMKIHPTBHXPF-TUJRSCDTSA-M 0.000 description 4
- 108010024957 Ascorbate Oxidase Proteins 0.000 description 3
- 102000004882 Lipase Human genes 0.000 description 3
- 108090001060 Lipase Proteins 0.000 description 3
- 239000004367 Lipase Substances 0.000 description 3
- 102000004316 Oxidoreductases Human genes 0.000 description 3
- 108090000854 Oxidoreductases Proteins 0.000 description 3
- 102000003992 Peroxidases Human genes 0.000 description 3
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 3
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 3
- 239000001166 ammonium sulphate Substances 0.000 description 3
- 235000011130 ammonium sulphate Nutrition 0.000 description 3
- 238000011534 incubation Methods 0.000 description 3
- 235000019421 lipase Nutrition 0.000 description 3
- 230000007774 longterm Effects 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 239000002609 medium Substances 0.000 description 3
- 108040007629 peroxidase activity proteins Proteins 0.000 description 3
- 239000000276 potassium ferrocyanide Substances 0.000 description 3
- YDBHVMTTYXWHLI-UHFFFAOYSA-N 2,4,6-tribromo-3-hydroxybenzoic acid Chemical compound OC(=O)C1=C(Br)C=C(Br)C(O)=C1Br YDBHVMTTYXWHLI-UHFFFAOYSA-N 0.000 description 2
- 241000193830 Bacillus <bacterium> Species 0.000 description 2
- 108010077078 Creatinase Proteins 0.000 description 2
- 108010066906 Creatininase Proteins 0.000 description 2
- ZGTMUACCHSMWAC-UHFFFAOYSA-L EDTA disodium salt (anhydrous) Chemical compound [Na+].[Na+].OC(=O)CN(CC([O-])=O)CCN(CC(O)=O)CC([O-])=O ZGTMUACCHSMWAC-UHFFFAOYSA-L 0.000 description 2
- FFDGPVCHZBVARC-UHFFFAOYSA-N N,N-dimethylglycine Chemical compound CN(C)CC(O)=O FFDGPVCHZBVARC-UHFFFAOYSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 239000000549 coloured material Substances 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000035484 reaction time Effects 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 239000000758 substrate Substances 0.000 description 2
- XOGGUFAVLNCTRS-UHFFFAOYSA-N tetrapotassium;iron(2+);hexacyanide Chemical compound [K+].[K+].[K+].[K+].[Fe+2].N#[C-].N#[C-].N#[C-].N#[C-].N#[C-].N#[C-] XOGGUFAVLNCTRS-UHFFFAOYSA-N 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 108090000531 Amidohydrolases Proteins 0.000 description 1
- 102000004092 Amidohydrolases Human genes 0.000 description 1
- 241000186073 Arthrobacter sp. Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 241000186249 Corynebacterium sp. Species 0.000 description 1
- 241000723247 Cylindrocarpon Species 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 101710084378 Lipase 2 Proteins 0.000 description 1
- 102000016943 Muramidase Human genes 0.000 description 1
- 108010014251 Muramidase Proteins 0.000 description 1
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 101710132589 Peroxidase 2 Proteins 0.000 description 1
- 229920002873 Polyethylenimine Polymers 0.000 description 1
- 229920005654 Sephadex Polymers 0.000 description 1
- 239000012507 Sephadex™ Substances 0.000 description 1
- 229920002684 Sepharose Polymers 0.000 description 1
- 241000122973 Stenotrophomonas maltophilia Species 0.000 description 1
- 125000000217 alkyl group Chemical group 0.000 description 1
- 150000001450 anions Chemical class 0.000 description 1
- 239000012736 aqueous medium Substances 0.000 description 1
- -1 aralkyl alcohols Chemical class 0.000 description 1
- 239000003613 bile acid Substances 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- CLEJXNXDIYBJSQ-UHFFFAOYSA-L calcium;dichloride;heptahydrate Chemical compound O.O.O.O.O.O.O.[Cl-].[Cl-].[Ca+2] CLEJXNXDIYBJSQ-UHFFFAOYSA-L 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 238000005352 clarification Methods 0.000 description 1
- 238000003759 clinical diagnosis Methods 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 108700003601 dimethylglycine Proteins 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 229960000274 lysozyme Drugs 0.000 description 1
- 235000010335 lysozyme Nutrition 0.000 description 1
- 239000004325 lysozyme Substances 0.000 description 1
- WRUGWIBCXHJTDG-UHFFFAOYSA-L magnesium sulfate heptahydrate Chemical compound O.O.O.O.O.O.O.[Mg+2].[O-]S([O-])(=O)=O WRUGWIBCXHJTDG-UHFFFAOYSA-L 0.000 description 1
- 239000002808 molecular sieve Substances 0.000 description 1
- 229940078490 n,n-dimethylglycine Drugs 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 229930027945 nicotinamide-adenine dinucleotide Natural products 0.000 description 1
- BOPGDPNILDQYTO-NNYOXOHSSA-N nicotinamide-adenine dinucleotide Chemical compound C1=CCC(C(=O)N)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OC[C@@H]2[C@H]([C@@H](O)[C@@H](O2)N2C3=NC=NC(N)=C3N=C2)O)O1 BOPGDPNILDQYTO-NNYOXOHSSA-N 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N phenol group Chemical group C1(=CC=CC=C1)O ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 1
- 210000002381 plasma Anatomy 0.000 description 1
- XAEFZNCEHLXOMS-UHFFFAOYSA-M potassium benzoate Chemical compound [K+].[O-]C(=O)C1=CC=CC=C1 XAEFZNCEHLXOMS-UHFFFAOYSA-M 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- URGAHOPLAPQHLN-UHFFFAOYSA-N sodium aluminosilicate Chemical compound [Na+].[Al+3].[O-][Si]([O-])=O.[O-][Si]([O-])=O URGAHOPLAPQHLN-UHFFFAOYSA-N 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
- C12N9/0012—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7)
- C12N9/0026—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7) acting on CH-NH groups of donors (1.5)
- C12N9/0032—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7) acting on CH-NH groups of donors (1.5) with oxygen as acceptor (1.5.3)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/26—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S435/00—Chemistry: molecular biology and microbiology
- Y10S435/8215—Microorganisms
- Y10S435/822—Microorganisms using bacteria or actinomycetales
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S435/00—Chemistry: molecular biology and microbiology
- Y10S435/8215—Microorganisms
- Y10S435/822—Microorganisms using bacteria or actinomycetales
- Y10S435/841—Chainia
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S435/00—Chemistry: molecular biology and microbiology
- Y10S435/8215—Microorganisms
- Y10S435/822—Microorganisms using bacteria or actinomycetales
- Y10S435/886—Streptomyces
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S435/00—Chemistry: molecular biology and microbiology
- Y10S435/8215—Microorganisms
- Y10S435/822—Microorganisms using bacteria or actinomycetales
- Y10S435/908—Streptovirticillium
Definitions
- the present invention is concerned with a new sarcosine oxidase with improved stability in comparison with known sarcosine oxidases, especially in detergent-containing analysis reagents.
- Sarcosine oxidases (E.C. 1.5.3.1) can be used, inter alia, for the enzymatic determination of sarcosine, creatine and creatinine, the enzymatic determination of creatinine in serum, plasma or urine being of especial importance in clinical diagnosis.
- creatinine amidohydrolase E.C. 3.5.2.10
- creatine amidinohydrolase E.C. 3.5.3.3
- sarcosine oxidase hydrogen peroxide is finally formed from creatinine in the stoichiometric ratio of 1:1 and this can be determined colorimetrically in a simple way.
- the stability of the chromogenic substances in the neutral, aqueous medium employed is also better than that of NADH, the indicator used in UV tests.
- the creatinine determination according to Jaffe Hoppe-Seyler's Z. Physiol. Chem., 10, 391/1886
- this process also offers a substantially greater specificity and thus an improved diagnostic dependability.
- the use of corrosive, strongly alkaline reagents is thereby also avoided.
- sarcosine oxidase for example for the determination of creatinine, requires that, in the reagent ready for use, it is sufficiently storage-stable for at least several days at 0° to 25° C. and, furthermore, in the case of carrying out the measurement even at elevated temperatures (30° to 37° C.), no significant loss of activity occurs over the minimum reaction time necessary.
- enzymatic analysis reagents preferably also contain so called clarification systems, most of which consist of a combination of lipases with non-ionic detergents (polyoxyethylated alkyl or aralkyl alcohols) and salts of bile acids, such as sodium cholate, as solubilising agent which permits the disturbance-free optical measurement even of strongly lipaemic samples.
- non-ionic detergents polyoxyethylated alkyl or aralkyl alcohols
- salts of bile acids such as sodium cholate
- the sarcosine oxidase has appropriate enzymatic properties, for example a low Michaelis constant for sarcosine and a high maximum reaction rate, since, due to these properties, there is essentially co-determined the reaction time in the case of sarcosine, creatine and creatinine determinations. Since the enzymatic determination of, for example, creatinine is also to be capable of being carried out at higher temperatures (37° C.) and in the presence of detergents and solubilisers, appropriate enzymatic properties are of considerable importance in the case of these stressing ambient conditions.
- a sarcosine oxidase obtainable from Streptomycetaceae which, at 25° C. in 0.15 mol/liter potassium phosphate (pH 7.9) and in the presence of surface-active substances, still shows after 2 days an activity of at least 40% of the initial activity.
- the sarcosine oxidase according to the present invention possesses for sarcosine a K M value of 2 to 4 mmol/liter.
- the K M values of the known sarcosine oxidases which are sufficiently stable for these determinations are, under these conditions, about 16 to 20 mmol/liter.
- the enzyme according to the present invention is found in all species of the family Streptomycetacea (The Prokaryotes, Vol. II (1981), 2028), for example in Chainia purpurogena DSM 43 156, Chainia ochraceae DSM 43 155, Streptomyces flocculus DSM 40 327, Streptoverticillium sp. DSM 40 237) and Kitasatoa purpurea DSM 43 362.
- the enzyme of the present invention is composed of four different sub-units and its molecular weight is about 170 kD.
- the enzyme is stable in the pH range of 6 to 9 and at temperatures below 40° C. At 50° C., it is inactivated within 15 minutes.
- the optimum temperature of the reaction is about 37° C. and the pH optimum is pH 8.0.
- the high substrate specifically is shown by the very low conversion of substrate analogues; thus, the conversion rate of, for example, N,N-dimethylglycine is only 1% of that of the sarcosine-specific reaction.
- the K M values for sarcosine (phosphate buffer; TES buffer), measured at 25° C. in various stressing reagents, are 2 to 3 mmol/liter.
- the V max is about 6 U/mg. protein.
- Table I gives, for various preparations of the enzyme according to the present invention, the K M values for sarcosine measured at 25° and at 37° C. For comparison, there are given the corresponding values for the known Bacillus enzyme.
- the enzyme according to the present invention has a superior stability in a detergent-containing medium at 37° C. which is shown not only by the crude extract supernatant but also by the purified enzyme.
- Table II shows the stability of the enzyme according to the present invention in comparison with known sarcosine oxidases.
- Table III shows the long-term stability at 25° C. of the enzyme according to the present invention and of the Bacillus sp. enzyme.
- composition as for stressing agent b but without the addition of the chromogenic colour system. Besides 0.15 mole potassium phosphate (pH 7.9), 0.1 mole TES/KOH is also used.
- the enzyme according to the present invention makes possible a substantially quicker carrying out of the enzymatic determination of sarcosine, creatine or creatinine. It has a very substantially better storage stability at 0° to 25° C. and , over the incubation interval, is, in the case of sarcosine, creatine or creatinine determinations at 37° C., substantially more stable than most of the known sarcosine oxidases.
- the organism was cultured in a complex medium of the following composition in a shaking flask: 5 g. yeast extract, 3 g. peptone (tryptic digested), 2 g. sodium chloride, 0.24 g. magnesium sulphate heptahydrate, 0.014 g. calcium chloride heptahydrate, 2 g. glucose, 10 g. sarcosine and 1 liter water (pH 7.0).
- the eluate was adjusted with ammonium sulphate to a concentration of 0.6 mole/liter and the enzyme was bound to phenyl-Sepharose and chromatographed with drecreasing ammonium sulphate gradients (above phosphate buffer).
- the eluates with over 4 U/mg. of protein were adjusted with ammonium sulphate to a concentration of up to 2.4 mole/liter.
- the precipitate was taken up in 0.1 mole/liter phosphate buffer and the sarcosine oxidase further purified by passage over a molecular sieve (Sephacryl-S-200, Pharmacia).
- the purified enzyme obtained had a specific activity of 5.5 U/mg. protein.
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- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Molecular Biology (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biomedical Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Immunology (AREA)
- Analytical Chemistry (AREA)
- Enzymes And Modification Thereof (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Detergent Compositions (AREA)
- Cosmetics (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE3519218 | 1985-05-29 | ||
| DE19853519218 DE3519218A1 (de) | 1985-05-29 | 1985-05-29 | H(pfeil abwaerts)2(pfeil abwaerts)o(pfeil abwaerts)2(pfeil abwaerts)-bildende sarcosinoxidase, ihre herstellung und verwendung |
Related Child Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| US07/153,976 Division US4845029A (en) | 1985-05-29 | 1988-02-09 | Method for the determination of sarcosine creatine or creatinine |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| US4743549A true US4743549A (en) | 1988-05-10 |
Family
ID=6271899
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| US06/868,262 Expired - Lifetime US4743549A (en) | 1985-05-29 | 1986-05-28 | Hydrogen peroxide-forming sarcosine oxidase |
| US07/153,976 Expired - Lifetime US4845029A (en) | 1985-05-29 | 1988-02-09 | Method for the determination of sarcosine creatine or creatinine |
Family Applications After (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| US07/153,976 Expired - Lifetime US4845029A (en) | 1985-05-29 | 1988-02-09 | Method for the determination of sarcosine creatine or creatinine |
Country Status (12)
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4845029A (en) * | 1985-05-29 | 1989-07-04 | Boehringer Mannheim Gmbh | Method for the determination of sarcosine creatine or creatinine |
| US5091312A (en) * | 1987-02-27 | 1992-02-25 | Kobayashi Pharmaceutical Co., Ltd. | Process for the preparation of sarcosine oxidase |
| US5565330A (en) * | 1992-10-01 | 1996-10-15 | Eli Lilly And Company | Method for removing N-terminal dipeptides from precursor polypeptides with dipeptidylaminopeptidase from Dictyostelium discoideum |
| EP3415910A1 (en) | 2017-06-16 | 2018-12-19 | Prevention Medicals s.r.o. | A method of quantitative determination of sarcosine in a biological sample using anti-arcosine antibodies and peroxidase-active gold nanoparticles or quantum dots |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| ES2026900T3 (es) * | 1986-08-28 | 1992-05-16 | Union Carbide Canada Limited | Descongelacion para aviones y composiciones anticongelante. |
| US9804154B2 (en) | 2013-03-12 | 2017-10-31 | Epinex Diagnostics, Inc. | Rapid test for urine albumin and urine creatinine |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4216292A (en) * | 1977-10-04 | 1980-08-05 | Toyo Jozo Kabushiki Kaisha | Process for the production of sarcosine oxidase |
| JPS5692790A (en) * | 1979-12-26 | 1981-07-27 | Hideaki Yamada | Preparation of sarcosine oxidase |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE3326888A1 (de) * | 1983-07-26 | 1985-02-14 | Boehringer Mannheim Gmbh, 6800 Mannheim | Sarcosin-oxidase |
| JPS61162174A (ja) * | 1985-01-11 | 1986-07-22 | Noda Sangyo Kagaku Kenkyusho | 耐熱性ザルコシン・オキシダ−ゼｎ及びその製造法 |
| DE3519218A1 (de) * | 1985-05-29 | 1986-12-04 | Boehringer Mannheim Gmbh, 6800 Mannheim | H(pfeil abwaerts)2(pfeil abwaerts)o(pfeil abwaerts)2(pfeil abwaerts)-bildende sarcosinoxidase, ihre herstellung und verwendung |
-
1985
- 1985-05-29 DE DE19853519218 patent/DE3519218A1/de not_active Withdrawn
-
1986
- 1986-04-28 ZA ZA863143A patent/ZA863143B/xx unknown
- 1986-04-29 AU AU56794/86A patent/AU561098B2/en not_active Ceased
- 1986-04-30 CA CA000507950A patent/CA1277269C/en not_active Expired - Lifetime
- 1986-05-13 ES ES554907A patent/ES8800345A1/es not_active Expired
- 1986-05-27 DE DE8686107203T patent/DE3679455D1/de not_active Expired - Lifetime
- 1986-05-27 AT AT86107203T patent/ATE63944T1/de not_active IP Right Cessation
- 1986-05-27 EP EP86107203A patent/EP0205967B1/de not_active Expired - Lifetime
- 1986-05-27 JP JP61120358A patent/JPS61280271A/ja active Granted
- 1986-05-28 FI FI862258A patent/FI91645C/fi not_active IP Right Cessation
- 1986-05-28 KR KR1019860004175A patent/KR890004091B1/ko not_active Expired
- 1986-05-28 SU SU4027546A patent/SU1582993A3/ru active
- 1986-05-28 US US06/868,262 patent/US4743549A/en not_active Expired - Lifetime
-
1988
- 1988-02-09 US US07/153,976 patent/US4845029A/en not_active Expired - Lifetime
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4216292A (en) * | 1977-10-04 | 1980-08-05 | Toyo Jozo Kabushiki Kaisha | Process for the production of sarcosine oxidase |
| JPS5692790A (en) * | 1979-12-26 | 1981-07-27 | Hideaki Yamada | Preparation of sarcosine oxidase |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4845029A (en) * | 1985-05-29 | 1989-07-04 | Boehringer Mannheim Gmbh | Method for the determination of sarcosine creatine or creatinine |
| US5091312A (en) * | 1987-02-27 | 1992-02-25 | Kobayashi Pharmaceutical Co., Ltd. | Process for the preparation of sarcosine oxidase |
| US5565330A (en) * | 1992-10-01 | 1996-10-15 | Eli Lilly And Company | Method for removing N-terminal dipeptides from precursor polypeptides with dipeptidylaminopeptidase from Dictyostelium discoideum |
| US5565349A (en) * | 1992-10-01 | 1996-10-15 | Eli Lilly And Company | Dictyostelium dipeptidylaminopeptidase |
| EP3415910A1 (en) | 2017-06-16 | 2018-12-19 | Prevention Medicals s.r.o. | A method of quantitative determination of sarcosine in a biological sample using anti-arcosine antibodies and peroxidase-active gold nanoparticles or quantum dots |
Also Published As
| Publication number | Publication date |
|---|---|
| DE3679455D1 (de) | 1991-07-04 |
| JPS61280271A (ja) | 1986-12-10 |
| CA1277269C (en) | 1990-12-04 |
| JPH0414957B2 (enrdf_load_stackoverflow) | 1992-03-16 |
| AU5679486A (en) | 1986-12-04 |
| FI862258A0 (fi) | 1986-05-28 |
| EP0205967A3 (en) | 1988-07-27 |
| US4845029A (en) | 1989-07-04 |
| ES554907A0 (es) | 1987-11-16 |
| AU561098B2 (en) | 1987-04-30 |
| FI91645B (fi) | 1994-04-15 |
| FI91645C (fi) | 1994-07-25 |
| KR890004091B1 (ko) | 1989-10-20 |
| ZA863143B (en) | 1986-12-30 |
| DE3519218A1 (de) | 1986-12-04 |
| EP0205967A2 (de) | 1986-12-30 |
| FI862258L (fi) | 1986-11-30 |
| KR860009127A (ko) | 1986-12-20 |
| EP0205967B1 (de) | 1991-05-29 |
| ES8800345A1 (es) | 1987-11-16 |
| SU1582993A3 (ru) | 1990-07-30 |
| ATE63944T1 (de) | 1991-06-15 |
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